Restriction endonucleases are enzymes that occur naturally in certain unicellular microbes—mainly bacteria and archaea—and that function to protect those organisms from infections by viruses and other parasitic DNA elements. Restriction endonucleases bind to specific sequences of nucleotides (‘recognition sequence’) in double-stranded DNA molecules (dsDNA) and cleave the DNA, usually within or close to these sequences, disrupting the DNA and triggering its destruction. Restriction endonucleases usually occur with one or more companion enzymes termed modification methyltransferases. Methyltransferases bind to the same sequences in dsDNA as the restriction endonucleases they accompany, but instead of cleaving the DNA, they alter it by the addition of a methyl group to one of the bases within the sequence. This modification (‘methylation’) prevents the restriction endonuclease from productively recognizing that site thereafter, rendering the site resistant to cleavage. Methyltransferases function as cellular antagonists to the restriction endonucleases they accompany, protecting the cell's own DNA from destruction by its restriction endonucleases. Together, a restriction endonuclease and its companion modification methyltransferase(s) form a restriction-modification (R-M) system, an enzymatic partnership that accomplishes for microbes what the immune system accomplishes, in some respects, for multicellular organisms.
A large and varied class of restriction endonucleases has been classified as ‘Type II’ class of restriction endonucleases. These enzymes cleave DNA at defined positions, and when purified can be used to cut DNA molecules into precise fragments for gene cloning and analysis. The biochemical precision of Type II restriction endonucleases far exceeds anything achievable by chemical methods, making these enzymes the reagents sine qua non of molecular biology laboratories. In this capacity as molecular tools for gene dissection Type II restriction endonucleases have had a profound impact on the life sciences and medicine in the past 25 years, transforming the academic and commercial arenas, alike. Their utility has spurred a continuous search for new restriction endonucleases, and a large number have been found: today more than 250 Type II endonucleases are known, each possessing different DNA cleavage characteristics (Roberts, R. J. et al., Nucl. Acids. Res. 33:D230–-D232 (2005)). (Rebase, http://rebase.neb.com/rebase). The production and purification of these enzymes have also been improved by the cloning and overexpression of the genes that encode them, usually in the context of non-native host cells such as E. coli. 
Since the various restriction enzymes appear to perform similar biological roles, and share the biochemistry of causing dsDNA breaks, it might be thought that they would resemble one another in amino acid sequence closely. Experience shows this not to be true, however. Surprisingly, far from sharing significant amino acid similarity with one another, most enzymes appear unique, with their amino acid sequences resembling neither other restriction enzymes nor any other known kind of protein. Type II restriction endonucleases seem to have arisen independently of each other during evolution, for the most part, and to have done so hundreds of times, so that today's enzymes represent a heterogeneous collection rather than a discrete family descended from a common ancester. Restriction endonucleases are biochemically diverse in organization and action: some act as homodimers, some as monomers, others as heterodimers. Some bind symmetric sequences, others asymmetric sequences; some bind continuous sequences, others discontinuous sequences; some bind unique sequences, others multiple sequences. Some are accompanied by a single methyltransferase, others by two, and yet others by none at all. When two methyltransferases are present, sometimes they are separate proteins and at other times they are fused. The orders and orientations of restriction and modification genes vary, with all possible organizations occurring. Several kinds of methyltransferases exist, some methylating adenines, others methylating cytosines at the N-4 position, or at the 5 position). Usually there is no way of predicting, a priori, which modifications will block a particular restriction endonuclease, which kind(s) of methyltransferases(s) will accompany that restriction endonuclease in any specific instance, nor what their gene orders or orientations will be.
From the point of view of cloning a Type II restriction endonuclease, the great variability that exists among R-M systems means that, for experimental purposes, each is unique. Each enzyme is unique in amino acid sequence and catalytic behavior; each occurs in unique enzymatic association, adapted to unique microbial circumstances; and each presents the experimenter with a unique challenge. Sometimes a restriction endonuclease can be cloned and over-expressed in a straightforward manner but very often it cannot, and what works well for one enzyme may fail altogether for the next. Success with one is no guarantee of success with another.
Novel endonucleases provide opportunities for innovative genetic engineering.